Crispr Cas9 Genbio2

CRISPR CAS9
Genbio2
Presentation
WHAT IS CRISPR CAS9?
CRISPR-Cas9 is a site-
specific gene editing
technology.
It is a component of bacterial
immune systems that can cut
a target DNA then shutting
the targeted gene off.
WHAT IS CRISPR CAS9?
CRISPR-Cas9 edits genes
and then harnessing natural
DNA repair processes to
modify the gene in the
desired manner.
CRISPR HISTORY:
In 1987 Atsuo Nakata and
his team of researchers
from Osaka University in
Japan first reported the
presence of CRISPR
CRISPR HISTORY:
In the mid 2000s, the functionality
and importance of CRISPR was first
realized in prokaryotes. As it turns out
CRISPR system is a part of
prokaryotic adaptive immunity, which
protects these prokaryotes from
attack by viral DNA, bacteriophages
and plasmids.
adaptive immunity
refers to that
immunity that an
organism aquires
after exposure to an
antigen or
vaccination.
CRISPR STANDS FOR:
CRISPR STANDS FOR:
Abreviated as CRISPR in
Escherichia coli genome, this
refer to short repeated
sequences of DNA nucleotides
found within the genome of
prokaryotes.
DEVELOPERS OF CRISPR CAS9 & WHY IT IS
MADE:
Microbiologist Biochemist
Emmanuelle Jennifer Doudna
Charpentier
They were first to propose that the bacterial CRISPR-
Cas9 system could be used as a programmable toolkit
for genome editing in humans and other animal species
DEVELOPERS OF CRISPR CAS9 & WHY IT IS
MADE:
Microbiologist Biochemist
Emmanuelle Jennifer Doudna
Charpentier
and they eventually recieve the noble prize in chemistry
for their work, in 2020.
How does CRISPR Cas9
work?
We first have to understand the function of
CRISPR in procaryotes, because
understanding the mechanism of its natural
function will be necessary in order to
understand the way it is exploited to achieve
genome editing capabilities in humans and
other organisms.
Exampl
e:
Exampl
e:
Once the viral DNA is injected

into the cell, a section of it can
take into the bacterial genome,
and will be inserted between the
repeated palindromic sequence.
Exampl
e:
This will now be called spacers. So

here we can see three different
spacers, potentially from three
different viruses, sandwiched in
Exampl
e:
Then we have now our CRISPR array,

this will undergo transcription to form
CRISPR RNA/crRNA and the longer
strand is called pre-crRNA.
Exampl
e:
Ca
sr
efe
r st
o:
Then the protein cas9 gets involved,

and as we know nucleases are
enzymes that are capable in
splitting/cutting the DNA at specific
Exampl
e:
Exampl
e: Along with cas9, there are also
a molecules of tracrRNA that is
complementary or can pair to
the palindromic repeat
Then another enzyme called
Exampl ribonuclease three (RNaseIII) wil
e: cut the strand in between,

leaving us with individual crRNA
which we can call effector
complexes
effector comlexes
Exampl
e:
if the effector complexes

encounters a section of
viral DNA that has a
sequence which is
complementary to this
crRNA the nuclease
Exampl
e:
and if it recognizes a short

sequence unique to the
viral genome called a
protospacer-adjacent motif
(PAM), then it will snip/cut
both strands of the DNA
Exampl
e:
in doing so, it will

neutralize the virus,
because its genome can no
longer be transcribed
properly to create more
viral particles, so infection
So that gives
us a
reasonable
understanding
of how CRISPR
is employed as
a natural
defense in
prokaryotes.
So how can a
genome
editing be
achieved using
this method?
in bacteria
crRNA + trcrRNA=
sgRNA
if the sgRNA complexes
with cas9, this two
component system will
be able to cleave DNA,
just as the three
component in bacteria
what this mean is it was
possible to determine 20
base pairs as a target for
editing, and all that has to
be done is to synthesize the
appropriate sgRNA with the
complementary sequence
and insert that into a cell
along with the cas9 protein
which have been sourced
from streptococcus
pyogenes.
The complex will form, read
the DNA until it finds the
appropriate sequence along
with a PAM sequence,
binding will occur, and DNA
will be cleaved at precisely
the desired location.
Cas9 has two domains,
and each one will snip one
of the DNA strands.
The cleaved dsDNA can
undergo repair via two
routes. Either by homology-
directed repair, or by non-
homologous end joining
The NHEJ pathway repairs double-strand
breaks in DNA by directly ligating
without the need for a homologous
template, which means a DNA strand
with similar sequence that can act as a
The NHEJ mechanism can also introduc
insertion or deletion of specific
sequences at the joining ends, thus
creating what are referred to as indels.
Indels are DNA strands with
either an insertion or
deletion of nucleotide
sequences. Thus, NHEJ
produces DNA strands with
non-uniformity in size.
The other route of repair,
the HDR pathway, is
commonly found in
bacterial and archaeal cells,
while the NHEJ pathway we
just discussed is more
common in a eukaryotic
The HDR process uses a
homologous DNA
template. The
homologous DNA
template has homology to
the adjacent sequences
surrounding the site of
cleavage to incorporate
new DNA fragments.
The template guides the
repair process, and lowers
the possibility of errors.
Since there is no insertion
or deletion of nucleotide
sequences, the HDR
pathway maintains
uniformity in the size of the
resulting dsDNA, unlike
NHEJ.
So that covers
the
mechanism of
CRISPR Cas9
as gene
editing tool.
APPLICATION OF CRISPR CAS9 IN
MEDICAL FIELD
MEDICAL FIELD
MEDICAL FIELD
THANK YOU!!!!
REFERENCES:
https://www.youtube.com/watch?
v=IiPL5HgPehs&pp=ygUMY3Jpc3ByIGNhcyA5
https://www.youtube.com/watch?v=2pp17E4E-
O8&pp=ygUMY3Jpc3ByIGNhcyA5

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CRISPR CAS9

Once the viral DNA is injected

This will now be called spacers. So

Then we have now our CRISPR array,

Then the protein cas9 gets involved,

e: cut the strand in between,

if the effector complexes

and if it recognizes a short

in doing so, it will

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